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Abstract

Background

Understanding how bone (BM), lean (LM) and fat mass (FM) develop through childhood,
puberty and adolescence is vital since it holds key information regarding current
and future health. Our study aimed to determine how BM, LM and FM track from prepuberty
to early adulthood in girls and what factors are associated with intra- and inter-individual
variation in these three tissues.

Methods

The study was a 7-year longitudinal cohort study. BM, LM and FM measured using dual-energy
X-ray absorptiometry, self-reported dietary information, leisure time physical activity
(LTPA) and other factors were assessed one to eight times in 396 girls aged 10 to
13 years (baseline), and in 255 mothers once.

Results

The location of a girl's BM, LM and FM in the lower, middle or upper part of the sample
distribution was established before puberty and tracked in its percentile of origin
over 7 years (r = 0.72 for BM, r = 0.61 for LM, and r = 0.65 for FM all p < 0.001 first vs. last measurements' ranking). Seventy-three percent of those in the
lowest quartile for BM and 69% for LM, and 79% of those in the highest quartile for
FM at baseline remained in their quartile at 7-year follow-up. Heritability was estimated
to contribute 69% of the total variance of the BM, 50% of the LM, and 57% of the FM.
Besides body size, diet index (explaining 9% of variance), breast feeding duration
(6%) and mother's BM (9%) predicted high BM. Diet index and high LTPA predicted high
LM (24% and 14%, respectively), and low FM (25% and 12%, respectively), and low level
of parental education predicted high FM (4%).

Conclusion

Individual levels of BM, LM and FM are established before puberty and track in a trait-specific
manner until early adulthood. Girls who are prone to develop low BM and LM and high
FM in adulthood can be identified in prepuberty. The developments of three components
of body composition are inter-related during growth. BM was the most heritable trait
while LM the most environmentally modifiable. Diet and physical activity played an
important role in increasing LM and preventing the accumulation of excessive FM.

Background

The biology of bone, muscle and fat tissues and their associated disorders such as
obesity, osteoporosis and sarcopenia are closely inter-related and share several common
origins, including genetic and environmental factors [1-5]. Skeletal muscle, acting in conjunction with energy expenditure and related hormone
regulation, is one of the major tissues responsible for metabolic status [4-6]. As muscle and adipose tissue are closely linked anatomically, biologically and pathologically,
study of the interrelationship between these two tissues is of great importance in
understanding the pathogenesis and development of diseases related to obesity and
physical activity/inactivity [6-8]. Furthermore, leptin, an adipocyte-derived hormone, is a major regulator of bone
remodelling [9]. A recent finding in an animal study showed that bone exerts an endocrine regulation
of energy homeostasis [2]. Together these studies emphasized that when studying the development of body composition,
bone mass (BM), lean mass (LM), and fat mass (FM) should be considered together.

Changes in BM, LM and FM during puberty are indicators of metabolic processes, and
thus hold pertinent information regarding current and future health [10]. Although it is well known that the components of body composition track, especially
from adolescence onwards [10], there is a paucity of information on how bone, muscle and fat tissues change during
the rapid phase of adolescent growth, and when non-optimal body composition characteristics
can be detected.

Early identification of the risk factors for future diseases is the first step towards
effective prevention strategies. In order to set up cost-effective programmes promoting
healthy growth in body composition, more attention should be paid to what factors
contribute to intra- and inter-individual variation in BM, LM and FM during adolescent
growth, and in what ways.

In this longitudinal study, we examined the development of three components of body
composition during growth and attempted to answer the following questions: i) Do BM,
LM and FM track, and if so, how are they inter-related from prepuberty to early adulthood?
ii) Can girls who are prone to develop low BM and LM and high FM in adulthood be identified
in prepuberty? iii) What factors are associated with intra- and inter-individual variation
in these three components of body composition?

Methods

Study population

The subjects were first contacted via class teachers teaching grades 4 to 6 (age 9
to 13 years old) in 61 schools in the city of Jyväskylä and its surroundings in Central
Finland (96% of all the schools in these areas). Briefly, of the eligible subjects,
396 girls participated in the laboratory tests one to eight times during a maximum
period of 8 years (mean duration of total follow-up was 7.5 years and mean age at
last follow-up was 18.3 years; n = 396 at baseline, n = 208 at 6-month, n = 201 at 12-month, n = 191 at 18-month, n = 220 at 24-month, n = 88 at 36-month, n = 61 at 48-month, and n = 236 at 84-month follow-up). Of the 396 girls, 258 participated in a calcium and
vitamin D intervention during the first 2 years [11]. Although no intervention effects on body composition were found, whether or not
they were in the intervention group was taken into account in the present analysis.
The study protocol was approved by the ethical committee of the University of Jyväskylä,
the Central Finland health care district, and the Finnish National Agency of Medicines.
Informed consent was given by all subjects and their parents prior to the assessments.

In addition, 255 mothers (age 32 to 58.9 years), 79 sisters (age 10 to 31 years),
and 41 brothers (age 9 to 31 years) participated in the same study procedures as the
girls. Among mothers, data on 144 premenopausal mothers (mean age 45.0, range 32 to
54 years) were used for comparison with their daughters at age 18 and to estimate
the maternal contribution to BM, FM, and LM. The sibling data were used in estimation
of the heritability in the broad sense. The participants provided their written consent
in accordance with the guidelines laid down by the ethical committees.

Background information

All the information on all participants was collected and laboratory tests performed
within two weeks' period during the same month of the year throughout the 7-year follow-up
to avoid seasonal effects.

Lifestyle and behavioural characteristics as well as medical history were collected
via a self-administered questionnaire. Girls under age 15 filled in the questionnaire
with their parents' assistance, and all the questionnaires were checked by a study
nurse. Breast feeding was expressed in months. The socio-economic factors included
the highest level of parental education, how wealthy the family is, and whether the
girl was living with both parents or a single parent (see Additional files 1 and 2). Birth crown-heel length and birth weight were obtained from the girl's growth chart.

Dietary information was obtained from a food-intake diary kept for three days (two
ordinary school days and one weekend day, see Additional file 3) as described elsewhere [12]. The food records contained time of eating, items and portion sizes. The families
were given written instructions and an example of how to record food consumption with
a help of portion guidebook. Details of all foods and drinks including the type and
commercial brand name were filled in the records. In addition, the type of diet (if
restricted) was requested in a separate question. During the coding process the food
records were checked by the nutritional students. Dietary intakes were analyzed using
Micro-Nutrica software (version 2.5), Finland. The program has been validated and
gives a reasonably good estimate of the intake of energy and most nutrients compared
with chemical analyses of the diet [13]. For this report, on the basis of earlier studies [14-17], the nutrients related to body composition were chosen to compute a dietary intake
index, including intakes of protein (g/day), calcium (Ca) (mg/day), potassium (K)
(mg/day), phosphorus (P) (mg/day) and magnesium (Mg) (mg/day) (for more details, see
statistical methods).

A leisure time physical activity (LTPA) score was calculated on the basis of questionnaire
(see Additional files 1 and 2) [18] responses as follows: the metabolic equivalents of the girl's three favourite leisure
time exercises (outside school) during the previous six months × duration of each
exercise session × the weight-bearing condition of each exercise (for example, non-weight-bearing
= 1 and weight-bearing = 2, as weight-bearing exercise is important for bone development)
× times/week of participation of those exercises. The score was validated against
a 7-day physical activity diary and heart rate monitor for estimating energy expenditure
in a sub-group; Bland-Altman analysis showed an acceptable agreement.

Anthropometrical and maturation assessments

Body height (cm) and weight (kg) were measured using standardized protocols and body
mass index (BMI = kg × m-2) was calculated. Sexual development was determined according to Tanner grading system
by a nurse [19]. The age of menarche was defined as the first onset of menstrual bleeding and was
collected by questionnaire, retrospective phone call, and/or interview during a clinical
visit.

Body composition assessments

Bone mass (BM in kg), lean tissue mass (LM in kg), and fat mass (FM in kg) of the
whole body were assessed using a dual-energy X-ray absorptiometry (DXA Prodigy; GE
Lunar Corp., Madison, WI USA) at baseline, 24 months, 36 months, 48 months and 84
months. The coefficient of variation (CV%) of two repeated measurements on the same
day was on average 0.7% for BM, 1.0% for LM, and 2.2% for FM in this study.

Statistical analyses

All data were checked for normality using the Shapiro-Wilk's W-test in SPSS 15.0 for Windows. Age-specific means and standard deviations of the
anthropometric and body composition traits were calculated. To evaluate if BM, LM
and FM track differently from prepuberty to early adulthood, a hierarchical linear
model with random effects was employed to explore the growth patterns of height, weight,
BM, LM, and FM (n = 396 girls). In this model, the time relative to menarche (TRM) was entered as the
explanatory variable in the form of polynomial spline functions to explain the change
in these variables over time (MLwiN 2.02 software, Multiple Project, Institute of
Education, University of London, UK).

To determine how well the girls' BM, LM and FM tracked from baseline to young adult,
we compared the within- and between individual variances of these three tissues, adjusting
for TRM, compared the proportion of the initial values that remained in the same quartile
at the 7-year follow-up visit, and correlated the initial and final measurements during
the 7-year period (n = 101 girls). Further, comparison of the variances between mother-daughter pairs was
assessed using the paired t-test (n = 144). Spearman's rho correlation was used to evaluate the percentile position for
BM, FM, and LM between baseline and the 7-year follow-up.

We used sibling data to estimate the broad sense heritability of body composition
with a linear mixed-effects model (n = 456 individuals, including brothers and sisters of the girls) [20,21]. In addition, the correlation between daughter (at the age of 18 years) and mother
(premenopausal) was used to estimate the maternal effect on the girl's body composition.

To determine what factors were associated with the intra- and inter-individual variation
of BM, LM, and FM, we used a generalized estimating equations model of longitudinal
data (GEE) with repeated measurements at ages 11 and 18 years. In this report the
following predictors of BM, LM and FM of the girls were included in the GEE model:
crown-heel length and birth weight, body height and weight, mother's BM, LM and FM,
breast feeding, diet index (protein, Ca, K, P, and Mg), LTPA, level of parental education,
time of assessment, that is, the baseline at the mean age of 11 and follow-up at the
mean age of 18, whether the girl was in the intervention during the first 2 years,
and age at menarche.

The R2 of the GEE model was computed according to Hardin and Hilbe [22], and it can be interpreted as a similar measure of the proportion of the outcome
variance explained by the model in common linear regression. Increments to R2 for each regression coefficient were calculated by a formula introduced in Natarajan
et al [23]. It is similar to the squared partial correlation computed in linear regression analysis
for type III sum of squares and it indicates what proportion of variance explained
relates to a given covariate that is unrelated to the other covariates in the model.

The significance of each regression coefficient was tested by comparing the Wald-test
statistics for a single degree of freedom with the standard normal distribution. To
overcome the problem of collinear predictors, we noted that collinearity is a problem
related to the testing of the regression coefficients and not description of data,
and subsequently we used two procedures. We used a principal component of collinear
covariates measured only at baseline in the GEE model (birth size index = principle
component of crown-heel length and birth weight). For covariates that were measured
both at baseline and at follow-up, the collinear predictors (diet = Protein, Ca, Mg,
K, P; body size = height, weight) were tested as a group using the Wald test statistic
for appropriate degrees of freedom (for diet cluster df = 5, body size cluster df
= 2 [24]. R2 values represent variance explained by the cluster of diet variables, that is, diet
index). This was considered useful also from the perspective that individuals do not
consume individual nutrients, but combinations of them, so it is their combined explanatory
power and statistical significance that is meaningful in this context. For prediction
the degree of collinearity should be similar in the target population as observed
here [25]. We observed this was the case when comparing the baseline and the follow-up data.

Results

Growth patterns

The growth patterns of BM, LM, and FM, by TRM in the girls are presented in Figure
1. Mothers' BM, LM and FM are presented in Figure 1 for comparison with the values recorded for the girls between 18 and 20 years of
age. Height nearly reached the peak value at 3 years post-menarche. Girls at the age
of 18 years were similar in height to their mothers (166.1 ± 5.4 vs. 165.2 ± 5.9 cm).
In contrast, the body weight of the girls was 9 kg (87%) less than that of their mothers'
owing to 0.16 kg (6%) less BM, 3.7 kg (9%) less LM, and more importantly, 5.2 kg (21%)
less FM.

Figure 1.Growth curves (left panels) for whole body bone mass, lean mass, and fat mass in girls. The x-axis is the time relative to menarche (months). In the left panel, points and thin
linking lines indicate individual measurements and the thick line represents the best
fitting line estimated by a hierarchical linear model with random effects. In the
right panel, each error-box indicates an age group value (mean ± 1 SD shade area).
The error line represents the 95% confidence interval.

Within and between-individual trait variances

During the 7 years of growth, the within-individual trait variances were 14% for height,
18% for weight, 20% for BM, 38% for LM and 24% for FM of the between-individual variances
(Table 1, Figure 2). The magnitude of the variance of BM, LM and FM was similar in girls at the age
of 18 to that of their mothers in their 40s (Figure 2). The results were similar when we compared only the mother-daughter pairs.

Figure 2.Residuals of bone mass, lean mass, and fat mass in girls compared with their pre-menopausal
mothers. Each point indicates an individual value. All values are in kg. For girls the value
adjusted for time relative to menarche.

Tracking of traits

The growth trajectories of BM, LM and FM tracked fairly well from pre puberty to early
adulthood. The baseline rank percentile of BM, LM and FM correlated with the follow-up
rank percentile 7 years later (r = 0.72 for BM, r = 0.61 for LM and r = 0.65 for FM, all p < 0.001). When tracked for decile position, 77% of the individuals remained within
their starting decile for BM, 69% for LM, and 66% for FM. To illustrate the tracking
of traits, the girls were grouped into quartiles according to the baseline measurements
and the four groups increased in parallel (Figure 3). Seventy-nine percent of those in the highest quartile for FM at baseline remained
in the highest quartile 7 years later; 73% and 69% in the lowest quartile for BM and
LM respectively, remained in their initial quartile 7 years later (Table 2). The girls in the lowest quartile of the BM group matured earlier (age of menarche
13.0 years) than those in the other quartiles (all at age 3.4 years, p = 0.032, 0.027 and 0.01, respectively). Similar differences in age of menarche were
found in the LM quartiles (lowest vs. 50 to 75% quartile p = 0.05, and lowest vs. highest p = 0.024). No differences in age of menarche were found between the FM quartile groups.

Table 2. The proportion of the subjects in the lowest and highest quartile at both baseline
and 7-year follow-up

Inter-relationships among the traits

The changes in BM, LM and FM correlated positively with each other during the 7-year
period (BM with LM r = 0.789, BM with FM r = 0.529, LM with FM r = 0.378, all p < 0.001). Sixty-two percent of the variance in BM gain could be explained by the change
in LM, while 28% could be explained by the change in FM. There was a negative correlation
between TRM and percentage gain in BM (r = -0.537, p < 0.001) and LM (r = -0.622,
p < 0.001) but not FM (r = -0.153, p = 0.126).

Predictors

In the GEE model, we found that the predictors of BM, LM, and FM in the girls were
different (Table 3). The common predictor for high BM, LM, and FM was a high body size index (explaining
62%, 59% and 78% of the variance, respectively), and for high BM and LM and low FM
it was diet index (explaining 9%, 24% and 25% of the variance, respectively); for
high BM the other predictors were high BM in the girl's mothers (9%), long duration
of breast feeding (6%) and low birth size (3%); for high LM another predictor was
high LTPA (14%); and for high FM low LTPA (12%). The time-dependent predictors assessed
at follow-up more strongly predicted BM, LM and FM than those assessed at baseline
(body size for BM, diet and LTPA for LM and FM).

Discussion

This study demonstrated that individual differences in BM, LM and FM are established
early in life: the variations in the traits seen in adults were already largely expressed
in prepubertal girls. The increase in BM was highest near menarche while high LM and
FM accumulation occurred much earlier before menarche. Further, we found that the
magnitude of the between-individual variance and the individual's relative positions
in the lower, middle or upper quartiles for BM, LM, and FM were established before
puberty. The ranking of one individual's trait at the baseline remained largely unchanged
7 years later.

The next question was whether these three tissues track differently. Previous studies
have reported that 69 to 92% of the variance of bone traits at maturity was accounted
for by variances at puberty [26,27]. Total FM, %FM and LM have also been reported to track from childhood to young adulthood
[28]. The tracking of total body FM and LM from early childhood into adulthood was low
but significant [29,30], whereas the tracking of FM and LM from post-puberty to adulthood was high [28]. In agreement with those studies our data showed that the tracking was stronger in
BM than LM and FM during a 7-year follow-up. Further, the correlation between daughters'
and their mothers' BM was stronger than that between their LM or FM. In addition,
heritability estimated using sibling data also showed that BM was the most heritable
trait while LM was the most environmentally modifiable.

In the light of the knowledge that these three tissues tracked differently, it would
be important to understand how they are related to each other during growth. We found
that the changes in BM, LM and FM correlated positively with each other during the
7-year period, suggesting that change in one tissue may have significant effects on
others. In particular, more than 60% of the gain in BM is accounted for by the gain
in LM.

Our data together with the results of earlier studies [31,32] suggest that it is possible to identify children who are prone to develop low BM
and LM and high FM. The importance of monitoring body composition during puberty resides
in the fact that many aspects of body composition during this period are predictive
of subsequent measures of these traits in adulthood (that is, body composition 'tracks')
[33,34] and, furthermore, are risk factors for many chronic diseases, for example cardiovascular
disease, diabetes mellitus, obesity and osteoporosis, in later life [35-38]. Even further, muscle weakness is a risk factor for mobility limitation and disability
and risks for falling among older populations [39-41]. LM has been widely used as a surrogate for muscle mass as well as muscle strength.
The intra- and inter-individual variability of these three tissues is partly accounted
for by a genetic component and partly by environmental factors. Tracking is influenced
by both heritable and non-heritable components, both of which need to be taken into
account when identifying those at risk for developing low BM and LM and high FM in
later life. Interestingly, the girls did not show an increment in LM after age 16.
However, by age 18, the girls already had reached 91% of their mother's LM values
but only 79% of their mothers FM. This stresses the importance of adolescence in the
development not only of BM and FM, but also of muscle mass. The findings on the inter-relationships
between the development of LM, BM and FM also suggest that the development of BM,
and even FM, during growth can be adjusted by increasing LM through interventions
such as increasing physical activity.

Of the modifiable factors, diet is one factor that has been shown to contribute importantly
to body composition during adolescence [42,43] and during aging [44]. In an earlier report we showed that supplements of calcium, dairy products and vitamin
D did not have a significant effect on bone mass accrual during puberty [11]. In the present study, the intervention did not have significant effects on body
composition 4 to 6 years after its conclusion. However, we found that our diet index
(a cluster of nutrients) was a strong predictor of BM, LM, and FM. This result clearly
underlines the importance of a habitual good diet. All of the nutrients chosen for
the model correlated with high milk consumption typical of the Finnish diet and have
been reported to have an effect on body composition [12]. In addition, we also found that longer duration of breast feeding was associated
with higher BM as has been reported earlier [45,46], and that a high level of parental education was associated with low FM. A higher
level of education may be associated with awareness of the importance of a good quality
of diet and with a higher income.

Being less physically active may contribute to the larger variation in FM among the
post-menarche female population by leading to high FM in early adulthood. Our data,
in accordance with data obtained from previous randomized controlled trials [47] and from studies in twins discordant for physical activity during adolescence [48], showed that high LTPA is an independent predictor for low gain in FM. As expected,
physical activity was associated with increased LM, as exercise increases muscle mass.
Increased use of muscles is linked logically to decreased FM.

The differences in the proportion of within-individual trait variances to between-individual
variances in the body composition traits indicates that bone trait variance in the
population is largely the result of individual differences in genetic makeup rather
than in life style factors [32]. However, this should not be interpreted to mean that an individual's position in
the population is immutably fixed. Studies have shown that long-term sports participation
during early adolescence results in greater accrual of BM [49-52]. Enhancement of LM seems to be a good predictor of BM accumulation. The effect of
physical activity on BM has been shown to be more pronounced in early puberty before
menarche [50,52-54], and bone may be more responsive to specific types of physical activity [55-57]. The effects of physical activity on bone are partly attained via muscular activity.
An optimal way of inducing bone to respond to physical activity would be to increase
its strength without much increasing its total weight. Hence our measure of total
bone mass may not be optimal for capturing all the clinically relevant properties
of bone as shown by peripheral quantitative computed tomography studies [51,58,59]. It may also be that BM reacts similarly to the increase in physical activity and
the gain in body weight during growth.

Although our research approach was rather wide ranging, it has a number of limitations.
It is difficult to establish the precise physiological mechanisms that drive the developmental
adaptation of human body composition from puberty to early adulthood on account of
the array of potentially confounding factors [60]. In our GEE model we did not include hormones, genotypes or fathers' data. Our supplementary
heritability estimation used only sibling data, which may affect the estimation of
the variance of these three tissues despite the similarity between our estimation
and the estimations given in other reports. The high drop-out rate in the follow-up
assessments was mainly due to a high proportion of girls going to universities or
professional schools and relocating at the end of the follow-up period. However, our
statistical methods were able to cope with this since hierarchical models allow inclusion
of data from every subject regardless of irregularly-spaced and missing data. The
strength of our study is that we were able to use 7-year longitudinal data with multiple
factors to assess the whole body composition, including bone, muscle and fat.

Conclusion

Our data suggest that prepubertal girls with low BM and LM and high FM are prone to
develop low values for peak bone and muscle mass and high fat mass in adulthood. A
habitual good quality diet is essential for the development of high bone and muscle
mass and normal fat mass during puberty. Physical activity is important in increasing
muscle mass and is an appropriate strategy for preventing high fat mass during the
adolescent growth period.

Competing interests

The study sponsors played no role in the design, methods, data management or analysis,
nor in the decision to publish. The corresponding author had full access to all the
data in the study and had final responsibility for the decision to submit for publication.
No authors hold any stocks or shares in an organization that may in any way gain or
lose financially from the publication of this manuscript. No authors have financial
or non-financial competing interests to declare in relation to this manuscript.

Authors' contributions

SC has full access to all of the data in the study and takes full responsibility for
the integrity of the data and for the accuracy of the data analysis. None of the authors
have financial or personal interest's affiliations with the sponsors of this research
effort. SC, FAT, HK, MA, UMK conceived of and designed the study. SC, EV, AL, SMC,
LX, MA were responsible for data acquisition. SC, EV, FAT, AL, TT, LX, SMC, HK, MA,
UMK analyzed and interpreted the data. SC, FAT, MA, UMK drafted the manuscript. SC,
EV, FAT, AL, TT, LX, SMC, HK, MA, UMK critically revised the manuscript for important
intellectual content. SC, EV, FAT, TT provided statistical expertise. SC obtained
funding. SC, EV, AL, TT, LX, SMC, HK, MA, UMK provided administrative, technical or
material support, and SC, MA, UMK supervised the project. All authors have read and
approved the final manuscript.

Acknowledgements

This study was supported by grants from the Academy of Finland, Finnish Ministry of
Education, PEURUNKA-Medical Rehabilitation Center, Candia Research Institute, France,
Jyväskylä City Science Park, EVO, Kuopio University, Jyväskylä Terveystalo, and American
Society of Bone and Mineral Research Bridge Funding.

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